We have isolated plasmids containing the Salmonella typhimurium origin of DNA replication (ori) and by construction of deletion derivative plasmids and cloning of restriction fragments of these plasmids, we were able to locate the functional origin to within a DNA fragment of 296 basepairs between the genes uncB and asn. The nucleotide sequence of the S. typhimurium ori region was determined and compared with the Escherichia coli ori sequence. In the 296 basepair fragment, 85.8 percent of the bases are conserved between the two species. A nearly equal number of transition and transversion type differences, with no insertions or deletions, occurs between the two bacterial origins, such that the relatively high percent adenine plus thymine of 59.5 percent is conserved. The high frequency of the sequence, GATC, which is the site of methylation under control of the dam gene, may explain, in part, why the bacterial ori greion appears to be so highly conserved.